Active suppression by regulatory T cells (Tregs) and T cell anergy are two important mechanisms for peripheral T cell tolerance. T cell receptor (TCR) signaling is critical for Treg function and induction of T cell anergy. However, TCR signaling can also activate T cells in the presence of a costimulatory signal. How TCR signaling is regulated to direct these distinct T cell fates/functions has been poorly understood. Diacylglycerol (DAG) kinases (DGKs) catalyze the conversion of DAG to phosphatidic acid through phosphorylation. We and others have recently demonstrated that DGK1 and 6, isoforms expressed in T cells, negatively control T cell activation by inhibiting TCR-induced activation of the DAG-RasGRP1-Ras-Erk1/2-AP1 cascade. Deficiency of either DGK1 or 6 causes T cells to be hyperresponsive to TCR stimulation and confers certain resistance to anergy induction. Our central hypothesis for this application is that DGK1 and 6 synergistically regulate DAG metabolism and play a pivotal role in T cell self-tolerance by promoting T cell anergy and Treg function. With strong support from preliminary data, we plan to test our hypothesis by pursuing three specific aims. In aim 1, we will determine how DGK1 and 6 synergistically control T cell anergy for self-tolerance. We will use already generated DGK1-/-6-/- mice as well as conditional DGK16 deficient mice being generated to test the hypothesis that DGK1 and 6 synergistically control the intrinsic property of T cells to promote anergy. In aim 2, we will investigate mechanisms by which DGK activity contributes to T cell anergy. In aim 3, we will elucidate the mechanisms by which DGK activity regulates Treg function. The proposed studies should significantly improve understanding of the mechanisms that regulate T cell anergy and Treg function, and identify strategies for modulating immune responses in order to treat autoimmune diseases. PUBLIC HEALTH RELEVANCE: The project proposed in this grant application aims to improve our understanding of the molecular mechanisms that control T cell anergy and regulatory T cell (Treg) function. We anticipate establishing and defining an important role of diacylglycerol kinase activity in T cell anergy, Treg function and self-tolerance to the liver. Studies in this grant proposal should also provide therapeutic targets to regulate T cell tolerance for autoimmune diseases.